Western blot analysis of Apolipoprotein A-1 was performed by loading the indicated amounts of a recombinant human Apo A-1-hIgG1Fc fusion protein (Product # 10686-H02H-50) or a recombinant mouse Apo A-1-hIgG1Fc fusion protein (Product # 50918-M02H-50), and 10ul of PageRuler Prestained Protein Ladder (Product # 26616) per well onto a Novex® 4-20% Tris-Glycine polyacrylamide gel (Product # WT4202). Proteins were transferred to a Nitrocellulose Membrane (Product # 88014) using the G2 Fast Blotter (Product # 62288), and blocked with 5% milk in TBST for at least 1 hour at room temperature. Apo A-1 was detected at ~50kD using an Apolipoprotein A-1 monoclonal antibody (Product # MA5-14668) at a dilution of 1:1000 in blocking buffer overnight at 4C on a rocking platform, followed by an HRP-conjugated goat anti-mouse IgG Fc-specific secondary antibody (Product # 31439) at a dilution of 1:40,000 for at least 30 minutes at room temperature. Chemiluminescent detection was performed using SuperSignal West Dura (Product # 34075).
|Tested species reactivity||Human|
|Host / Isotype||Mouse / IgG1|
|Immunogen||Purified human plasma Apo A-1|
|Storage buffer||0.01M potassium phosphate with 0.135M NaCl, 2.5mM KCl, 1mg/ml BSA|
|Contains||0.05% sodium azide|
|Storage Conditions||Maintain refrigerated at 2-8°C for up to 6 months. For long term storage store at -20°C|
|Tested Applications||Dilution *|
|Immunoprecipitation (IP)||Assay Dependent|
|Radioimmune Assays (RIA)||Assay Dependent|
|Western Blot (WB)||1:500-1:2000|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
By sandwich ELISA, MA5-14668 can be used as a detection antibody with Product # 710263 or # 701239 as a coating antibody, to generate a matched pair. Using these matched pairs, recombinant human Apo A-1, but not recombinant mouse Apo A-1, was detected. MA5-14668 can be used to detect Apo A-1 from serum samples. To increase sensitivity of sandwich ELISAs with MA5-14668, a biotinylated detection antibody followed by Streptavidin-HRP is recommended.
By Western blot, MA5-14668 detects recombinant human Apo A-1, but not recombinant mouse Apo A-1. MA5-14668 may not be as successful for detecting endogenous Apo A-1 by Western blot.
Product MA514668 is a smaller package size of MIA1402 (formerly sold as a Seradyn product).
This gene encodes apolipoprotein A-I, which is the major protein component of high density lipoprotein (HDL) in plasma. The protein promotes cholesterol efflux from tissues to the liver for excretion, and it is a cofactor for lecithin cholesterolacyltransferase (LCAT) which is responsible for the formation of most plasma cholesteryl esters. This gene is closely linked with two other apolipoprotein genes on chromosome 11. Defects in this gene are associated with HDL deficiencies, including Tangier disease, and with systemic non-neuropathic amyloidosis.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.